The present disclosure relates to a measurement apparatus and a measurement method.
A vibration spectral region that is important in considering application of vibrational spectroscopy is in the range of 300 cm−1 to 3600 cm−1 that is known as a molecular fingerprint region. As a method for measuring a vibration spectrum corresponding to the region having these wavenumbers, an infrared spectroscopic method and a Raman spectroscopic method are representative methods, and by using both measurement methods, complementary information relating to molecular vibration of a sample can be obtained. Here, in the case of a sample such as a biological sample that contains water as a main ingredient, a vibration spectrum caused by water is observed in the infrared spectroscopic method, and thus the Raman spectroscopic method is mostly used.
However, in analysis, examination, and diagnosis of a biological material, a Raman spectrum of the biological material generally includes many vibration spectra of molecular function groups and is accompanied with autofluorescence of the biological material, and thus the spectrum is broadened in a complicated manner and there are often difficulties in attribution of the functional groups. Furthermore, optical damage of the biological material relatively easily occurs, and therefore detection with high sensitivity has been demanded.
As non-linear Raman spectroscopic methods which are one kind of the Raman spectroscopic method, there are a coherent anti-Stokes Raman scattering (Coherent Anti-Stokes Raman Scattering; CARS) spectroscopic method and a stimulated Raman scattering (Stimulated Raman Scattering; SRS) spectroscopic method. Since the non-linear Raman spectroscopic methods described above have superiority in avoidance of autofluorescence of a sample, high sensitivity, and three-dimensional spatial resolution, application of the methods to microscopes has been remarkably developed.
On the other hand, since measurement targets of a biological sample are organic composite materials such as protein, fat, and water, spectra of organic molecules unique to functional groups are superimposed on each other in vibration spectroscopy such as general Raman spectroscopy, and thus there are many cases in which attribution of the spectrums is difficult.
Thus, in the CARS spectroscopic method, research and development have been conducted not only for measurement of a wavenumber spectrum of molecular vibration but also for measurement of a time domain, i.e., measurement of a relaxation time as a response of molecular vibration to an external stimulus of laser light. Vibration of a functional group corresponding to a specific spectrum is affected exactly by a surrounding environment of the functional group, i.e., interaction with a peripheral molecular, and a relaxation time thereof is changed in the range of several hundred fs (femtoseconds) to dozens of ps (picoseconds). In the general Raman spectroscopy (in other words, a wavenumber domain), a reciprocal of a spectrum width derived from uniform broadening of a spectrum corresponds to a relaxation time, but high spectrum resolution is required, and thus measurement is difficult as described above. On the other hand, measurement of a relaxation time neither requires spectrum resolution nor is sensitive to a peak position of a spectrum, and thus each measurement is an advantage.
However, in a relaxation time measurement method using the CARS, there is a problem in that decision of a relaxation time becomes difficult due to being affected by a non-resonant background.
Thus, as shown in Patent Literature 1 described below, a method for measuring a relaxation time of focused molecular vibration by using a stimulated Raman scattering (SRS) microspectroscopic method has been proposed.